Covalent post-translational modifications (PTMs) of epigenomic proteins contribute to their biological roles, and thus serve as carriers of epigenetic information from one cell generation to the next. Epigenetics means on top of or above genetics, and refers to external modifications to DNA and associated histones that turn genes “on” or “off.” These modifications do not change the DNA sequence, but instead, they affect how cells “read” genes. PTMs play key roles in the regulation of protein function, transcription, DNA replication, and repair of DNA damage (Lakshmaiah, K. C et al., J. Cancer Res. Ther. 2014, 10, 469-478).
The major events surrounding epigenetic control are focused on three modes of action: writers, readers, and erasers. The writers are responsible for adding a variety of PTM marks to histones which include, inter alia, acetylation which is catalyzed by histone acetyltransferases (HATs). Readers refer to the proteins that recognize and bind to these PTM marks thereby mediating their effects, and erasers encompass various enzymes such as the histone deacetylases (HDACs) that catalyze the removal of these marks. In the case of acetylated histone lysine residues, HDACs are responsible for catalyzing the hydrolysis of the acetyl mark to provide the unsubstituted lysine residue. The HDAC family consists of at present 18 enzymes which are classified into four subgroups according to their homology to the yeast family. HDAC1, 2, 3 and 8—categorized as class I HDACs according to their homology with yeast Rpd3—are characterized by ubiquitous expression and localization to the nucleus. Class II HDACs show tissue-specific expression and shuttle between the nucleus and cytoplasm. Homologous to yeast Hda1, these enzymes are subdivided in class IIa (HDAC4, 5, 7 and 9) and class IIb (HDAC6 and 10). HDAC11, the only member of the class IV subfamily, shows similarities to the catalytic domains of both class I and II enzymes. Class I, II, and IV HDACs require Zn2+ as a cofactor of the deacetylating activity and are also referred to as the conventional HDACs. The sirtuins 1-7 are dependent on nicotinamide adenine dinucleotide for their activity and form class III of the HDACs.
Pharmacologic manipulation of the enzymes involved in regulating protein PTMs, especially those tied to very specific PTM marks, holds tremendous possibilities in better understanding the workings of the cell. The discovery of selective small molecule modulators of these enzymes would provide chemical tools to better understand the role of these PTMs at the cellular level, but may also lead to important disease modifiers. Within the HDAC field, there exists a plethora of compounds that are able to block the deacetylase enzymes, and several have made their way to the marketplace for cancer therapy. The majority of these HDACIs, however, are not very isoform selective. Many of them inhibit across more than one class of HDAC enzymes and are thus labeled pan-selective. Of the various HDAC isoforms that appear to be promising therapeutic targets for treating human diseases such as cancer and certain CNS disorders, HDAC6 has emerged as a particularly attractive target, especially in view of the fact that HDAC6 knockout animals remain viable. HDAC6 has no apparent role in the PTM of histone proteins, but rather is involved in regulating the acetylation status of α-tubulin, HSP-90, cortactin, HSF-1, and other protein targets. This enzyme also plays a role in the recognition and clearance of polyubiquitinated misfolded proteins from the cell through aggresome formation. The development of HDAC6 selective compounds has recently been reviewed (Kalin, J. H. et al., J. Med. Chem. 2013, 56, 6297-6313). In general, HDACIs are comprised of three main motifs: a zinc binding group (ZBG), a cap group, and a linker that bridges the previous two (FIG. 1). A properly optimized cap group can improve both potency and selectivity, presumably through its ability to engage in appropriate contacts with residues on the enzyme surface.

Many HDACIs such as trichostatin A (TSA) and SAHA contain a hydroxamic acid function as ZBG (FIG. 1). Unfortunately, hydroxamates are in some cases metabolically unstable (short half-life), and their potent metal-chelating ability might lead to off-target activity at other zinc-containing enzymes (Flipo, M. et al., J. Med. Chem. 2009, 52, 6790-6802). In addition, many of the hydroxamic acid based inhibitors have been found to be Ames-positive, suggesting that these agents might present genotoxic effects. While several of the HDACIs on the market are Ames positive and cause chromosomal aberrations, these are being used only for cancer, wherein this undesired side effect can to a certain extent be tolerated in a disease considered to be life threatening. Certainly, for use in diseases that would require chronic, longer term use of an HDACI, it would be preferable to have compounds that are not Ames positive/genotoxic. However, even for cancer, it is known that use of genotoxic agents can lead to a genomic instability that may be transmitted to offspring in cases where the treated adults have children (Glen, C. D. et al., Proc. Natl. Acad. Sci. U.S.A. 2012, 109, 2984-2988). As such, there is a great need for the discovery of potent and selective HDACIs that bear alternative ZBGs or that are hydroxamates that for various reasons fail to show Ames activity. Our research has led to the discovery of hydroxamate based HDACIs that show high selectivity for the inhibition of HDAC6 but are not Ames active (Kozikowski, A. P. et al., J. Med. Chem. 2007, 50, 3054-3061).
In summary, extensive evidence supports a therapeutic role for HDACIs in the treatment of a variety of conditions and diseases, such as cancers and CNS diseases and degenerations. However, despite exhibiting overall beneficial effects, like beneficial neuroprotective effects, for example, HDACIs known to date have little specificity with regard to HDAC inhibition, and therefore inhibit all zinc-dependent histone deacetylases. It is still unknown which is (are) the salient HDAC(s) that mediate(s) neuroprotection when inhibited. Emerging evidence suggests that at least some of the HDAC isozymes are absolutely required for the maintenance and survival of neurons, e.g., HDAC1. Additionally, adverse side effect issues have been noted with nonspecific HDAC inhibition. Thus, the clinical efficacy of present-day nonspecific HDACIs for stroke, neurodegenerative disorders, neurological diseases, and other diseases and conditions ultimately may be limited. It is important therefore to design, synthesize, and test compounds capable of serving as potent, and preferably isozyme-selective, HDACIs that are able to ameliorate the effects of neurological disease, neurodegenerative disorder, traumatic brain injury, cancer, inflammation, malaria, autoimmune diseases, immunosuppressive therapy, and other conditions and diseases mediated by HDACs.
An important advance in the art would be the discovery of HDACIs, and particularly selective HDAC6 inhibitors, that are useful in the treatment of diseases wherein HDAC inhibition provides a benefit, such as cancers, neurological diseases, traumatic brain injury, neurodegenerative disorders and other peripheral neuropathies, stroke, hypertension, malaria, allograft rejection, rheumatoid arthritis, and inflammations. Accordingly, a significant need exists in the art for efficacious compounds, compositions, and methods useful in the treatment of such diseases, alone or in conjunction with other therapies used to treat these diseases and conditions. The present invention is directed to meeting this need.